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Overproduction, Purification and Characterization of Adenylate Deaminase from Aspergillus oryzae

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Abstract

Adenylate deaminase (AMPD, EC 3.5.4.6) is an aminohydrolase that widely used in the food and medicine industries. In this study, the gene encoding Aspergillus oryzae AMPD was cloned and expressed in Escherichia coli. Induction with 0.75 mM isopropyl β-d-l-thiogalactopyranoside resulted in an enzyme activity of 1773.9 U/mL. Recombinant AMPD was purified to electrophoretic homogeneity using nickel affinity chromatography, and its molecular weight was calculated as 78.6 kDa. Purified AMPD exhibited maximal activity at 35 °C, pH 6.0 and 30 mM K+, with apparent K m and V max values of 2.7 × 10−4 M and 77.5 μmol/mg/min under these conditions. HPLC revealed that recombinant AMPD could effectively catalyse the synthesis of inosine-5′-monophosphate (IMP) with minimal by-products, indicating high specificity and suggesting that it could prove useful for IMP production.

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Acknowledgments

This research was financially supported by Guangxi Natural Science Foundation (2015GXNSFBA139064), Fund of Guangxi Academy of Sciences (15YJ22SW05), the fund of Key Lab of Local Agricultural Product Depth Processing and Food Security Guarantee and the “Bagui Scholars Distinguished Professor” Special Project.

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Authors and Affiliations

  1. College of Light Industry and Food Engineering, Guangxi University, Nanning, 530004, China

    Shubo Li & Mouming Zhao

  2. College of Life Science and Technology, Guangxi University, Nanning, 530004, China

    Yi Qian, Yunlong Liang, Xinkuan Chen & Zongwen Pang

  3. National Engineering Research Center for Non-Food Biorefinery, Guangxi Academy of Sciences, Nanning, 530004, China

    Yuan Guo

  4. College of Life Science and Technology, Guangxi University, Nanning, 530005, China

    Zongwen Pang

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  1. Shubo Li
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  2. Yi Qian
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  3. Yunlong Liang
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  4. Xinkuan Chen
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  5. Mouming Zhao
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  6. Yuan Guo
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Correspondence to Yuan Guo or Zongwen Pang.

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Li, S., Qian, Y., Liang, Y. et al. Overproduction, Purification and Characterization of Adenylate Deaminase from Aspergillus oryzae . Appl Biochem Biotechnol 180, 1635–1643 (2016). https://doi.org/10.1007/s12010-016-2192-7

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  • DOI: https://doi.org/10.1007/s12010-016-2192-7

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